Structural implications of spectroscopic characterization of a putative zinc finger peptide from HIV-1 integrase
| Autor: | C J, Burke, G, Sanyal, M W, Bruner, J A, Ryan, R L, LaFemina, H L, Robbins, A S, Zeft, C R, Middaugh, M G, Cordingley |
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| Rok vydání: | 1992 |
| Předmět: |
Integrases
Protein Conformation Molecular Sequence Data Zinc Fingers Cobalt Hydrogen-Ion Concentration Recombinant Proteins Models Structural Kinetics Zinc Spectrometry Fluorescence Chlorides Spectrophotometry Zinc Compounds DNA Nucleotidyltransferases Chromatography Gel HIV-1 Mutagenesis Site-Directed Thermodynamics Amino Acid Sequence Cloning Molecular |
| Zdroj: | The Journal of biological chemistry. 267(14) |
| ISSN: | 0021-9258 |
| Popis: | The N-terminal domain of human immunodeficiency virus (HIV-1) integrase (IN) contains the sequence motif His-Xaa3-His-Xaa23-Cys-Xaa2-Cys, which is strongly conserved in all retroviral and retrotransposon IN proteins. This structural motif constitutes a putative zinc finger in which a metal ion may be coordinately bound by the His and Cys residues. A recombinant peptide, IN(1-55), composed of the N-terminal 55 amino acids of HIV-1 IN was expressed in Escherichia coli and purified. Utilizing a combination of techniques including UV-visible absorption, circular dichroism, Fourier transform infrared, and fluorescence spectroscopies, we have demonstrated that metal ions (Zn2+, Co2+, and Cd2+) are bound with equimolar stoichiometry by IN(1-55). The liganded peptide assumes a highly ordered structure with increased alpha-helical content and exhibits remarkable thermal stability. UV-visible difference spectra of the peptide-Co2+ complexes directly implicate thiols in metal coordination, and Co2+ d-d transitions in the visible range indicate that Co2+ is tetrahedrally coordinated. Mutant peptides containing conservative substitutions of one of the conserved His or either of the Cys residues displayed no significant Zn(2+)-induced conformational changes as monitored by CD and fluorescence spectra. We conclude that the N terminus of HIV-1 IN contains a metal-binding domain whose structure is stabilized by tetrahedral coordination of metal by histidines 12 and 16 and cysteines 40 and 43. A preliminary structural model for this zinc finger is presented. |
| Databáze: | OpenAIRE |
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